The invention pertains to lanthanide ion-ligand complexes, in particular to neodymium(III) ion (Nd3+), ytterbium(III) ion (Yb3+), or erbium(III) ion (Er3+) ligand complexes, to the use of said lanthanide ion-ligand complexes for the manufacture of a diagnostic kit, to a diagnostic kit comprising the same, and to a method of detecting an analyte in a matrix of biomedical interest.
The use of metal chelates as luminescent probes is well-known in the art, and the use of such probes for diagnostic purposes has recently been commercialized, while the state of the art of the technique has been reviewed by P. G. Sammes and G. Yahioglu in Natural Product Reports, Vol. 13, pp. 1-28 (1996). Europium and terbium ions (Eu(III) and Tb(III), respectively), both members of the rare-earth lanthanide metals, are very suitable as luminescent probes because of their long-lived luminescence, which allows for interference-free detection. By using a time delay between excitation pulse and detection optimized for the rare-earth ion, short-lived scatter and background luminescence from the matrix can be effectively removed. Since the absorption coefficients of rare-earth ions are extremely low (1-10 I.mole−1.cm−1) direct excitation is very inefficient. In general, excitation therefore requires the aid of a chromophore as sensitizer. For Tb(III) and Eu(III) ions, which emit in the visible part of the spectrum, sensitizers requiring UV excitation have to be applied as a result of energy constraints (the triplet state of the sensitizer should be at least 1000 cm−1 above the accepting state of the lanthanide ion). The other lanthanides in chelated forms in solution which exhibit luminescence in the visible part of the spectrum are gadolinium, samarium, and dysprosium ions. The other members of the lanthanide metals are considered to be unsuitable as luminescent probes because they have much smaller gaps between the excited states and the ground state. When used as a luminescent probe, the lanthanides are coordinated to form complexes with ligands, which complexes often are unstable in aqueous solutions.
Attempts have been made to obtain stable chelates of Eu(III) and Tb(III) ions, for instance, as disclosed in WO 96/00901 wherein complexes of Eu(III) and Tb(III) ions with polynuclear heterocyclic aromatic compounds are described. These chelate-lanthanide complexes may be applied as labels or probes for diagnostics, and are capable of intense luminescence in the visible region. A drawback to these Eu(III) and Tb(III) ion-chelates, however, is the need to sensitize them at a relatively short wavelength (UV or near-UV), which requires expensive lasers. Moreover, the excitation energy used regularly interferes with the biological material which is used for diagnostic purposes, which more often than not is sensitive to light of the UV region. Further it is of advantage to have a metal ion which exhibits a long lifetime in the excited state, which requires minimization of quenching and back-transfer.
In Russian patent applications SU 1340087 and SU 1621720 ytterbium porphyrin complexes have been disclosed, which are used to enhance the contrast coefficient in in vivo luminescent investigations of malignant tissues in animals. These complexes are not suitable as diagnostic assays for detection of specific analytes, because they do not contain an (immuno)-reactant for attachment to said analyte. Further, these complexes are hardly soluble and have poor stability in water.